The maintenance of immune tolerance is critical for the prevention of autoimmunity. A key cell that helps promote immune tolerance is the thymic medullary epithelial cell (mTEC). A critical subset of mTEC's expresses a wide array of tissue restricted antigens under the control of the Autoimmune Regulator (Aire) gene. Aire was identified as the defective gene in the autoimmune syndrome Autoimmune Polyglandular Syndrome Type 1 and highlights the critical importance of this gene and the mTEC cell population for the maintenance of tolerance. The Aire-expressing subset of mTEC's promotes self-tolerance by helping promote the display of TRA's to developing thymocytes for negative selection and perhaps the positive selection of regulatory T cells. Within the thymus, the mTEC compartment appears to be in a state of dynamic turnover, but what molecular pathways and cell populations that are involved in this turnover remain to be determined especially in adulthood after thymic development. In addition to their turnover, the role of mTEC's in the induction and selection of regulatory T cells (Treg's) in the thymus remains to be clearly delineated. Recently, we have developed a powerful set of genetic tools to help dissect and examine the turnover of mTEC's and their control of Treg selection including a cell ablation system and cell fate mapping system. Both systems can be temporally controlled and allow for testing cell turnover dynamics in the adult and neonatal settings. We hypothesize that the mTEC compartment is a dynamic compartment of cells that is regenerated by precursor cells that express Aire and that this cell compartment is also an important contributor to the selection of Treg's. To test these questions our specific aims are: 1) Determine the mechanism(s) of mTEC turnover and regeneration in adult mice, 2) Identify the role of mTECs in guiding regulatory T cell induction and development, and 3) Assess the functional consequences of Aire-expressing cell ablation on the development of autoimmunity. Taken together our studies will help determine what factors and cells contribute to the turnover and replacement of mTEC's, their potential role in the induction of Treg's, and whether the long term maintenance of mTEC's are critical for immune tolerance.